Published May 30, 2006
Research Roundup
D
Disordered domains can evolve quickly, given their structural freedom, and often bind several partners. As disordered
regions are commonly signaling and regulatory domains, more
disorder and more splicing might have contributed to the
emergence of cellular specialization. “With different splicing
in different cells,” says Dunker, “the signaling network becomes
radically altered.” Splicing out a piece of a disordered region
in BRCA1, for example, eliminates p53 binding.
Testing this evolutionary hypothesis, however, will
take time. To start, the group
would like to work out the
major signaling network differences between a pair of
similar but distinct cell types,
perhaps from a simple multicellular organism like the
sponge, and then determine
whether the changes relate
to alternative splicing within
disordered regions.
Reference: Romero, P.R., et al.
2006. Proc. Natl. Acad. Sci.
USA. 103:8390–8395.
Highly disordered regions are
more common in alternatively
spliced regions (left) than the rest of
the protein (right).
C a g e fo l d i n g
well-designed chaperone cage
helps proteins to fold quickly, say
Yun-Chi Tang, F. Ulrich Hartl,
Manajit Hayer-Hartl, and colleagues (Max
Planck Institute of Biochemistry, Martinsried,
Germany), probably by limiting the number
of possible folding intermediates.
Bacteria’s most well-studied chaperone
is the GroEL nano-cage, which encapsulates a folding substrate within its walls.
This cage was thought to be little more
than a way to isolate substrates to prevent
aggregation of slow-folding proteins. But
the new findings suggest an active role, as
folding rates inside the cage were up to
15-fold faster than in solution.
Folding is hastened by several cage features, including cage size. For small proteins, GroEL mutants with a smaller cavity
further accelerated folding, until a point at
which necessary rearrangements were spatially restricted. Confinement hastens folding by preventing those misfolded intermediates that would not fit within the cage.
“The number of possible conformations,”
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JCB • VOLUME 173 • NUMBER 6 • 2006
HAYER-HARTL/ELSEVIER
A
Decreasing GroEL cavity size (from left to right)
increases then decreases protein folding rate.
says Hartl, “is astronomically large. The
cage reduces it to a subfraction of that.”
For large proteins, both smaller and
larger cavities slowed folding. The cage
thus seems to be evolutionarily optimized
to suit its ∼250 in vivo substrates. “You
can improve folding rates for some with
GroEL mutations,” says Hayer-Hartl, “but
only at the expense of other substrates.”
Flexible, finger-like extensions at the
bottom of the cavity were necessary for
some proteins to fold quickly. The authors
speculate that these mildly hydrophobic
sequences gently massage misfolded
states, increasing the substrate’s fluidity
and easing rearrangements.
Some proteins also required clusters
of negative charges on the cavity wall for
optimal folding. Proteins most impeded by
the loss of these clusters were themselves
negatively charged, so perhaps the clusters
help by keeping these proteins from
sticking to the cage walls.
The authors would next like to identify intermediates in spontaneous and
GroEL-mediated folding events to see
just how the folding landscape changes
inside the cage.
Reference: Tang, Y.-C., et al. 2006. Cell.
125:903–914.
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isordered regions in proteins are prime real estate for
alternative splicing, say Pedro Romero, Keith Dunker
(Indiana University-Purdue University, Indianapolis, IN),
and colleagues find that d. The innate disorder allows protein
variation that in turn increases functional diversity.
Both splicing and disorder are more common in multicellular eukaryotes than in lower organisms. Dunker wondered whether this trend might be more than coincidence.
Structures are available for only five pairs of alternatively
spliced isoforms but, in three pairs, the regions present in
one splice form but absent in another are found within disordered regions.
To expand the dataset, the authors compared various
databases of disorder and of splicing. Indeed, alternatively
spliced regions were strongly biased toward disorder. Their
flexibility probably improves the odds that an addition or deletion will not impede folding and thus lead to aggregates.
Splicing in disordered regions might also increase functional diversity. Unlike structured domains, which use farflung residues to build a single functional unit, disordered
regions often use a compact and linear series of residues
to create a particular functional unit. “You get more bang
for your buck,” says Dunker. “Just splice out ten consecutive
residues, and a whole function is gone.”
DUNKER/NAS
Disorder for variety
Published May 30, 2006
Text by Nicole LeBrasseur
G e n e s p u l s e w i t h a c t iv i t y
Signaling from keratins
onathan Chubb, Robert Singer, and colleagues (Albert Einstein
College of Medicine, Bronx, NY) have devised a method to
view transcription inside individual cells. By removing the
effects of population averaging, the method reveals transcriptional
pulses of a eukaryotic developmental gene.
Transcription was visualized by inserting upstream stem loops into
an endogenous developmentally regulated Dictyostelium gene called
dscA. The nascent transcripts were then detected by a GFP fusion
protein that binds to the RNA stem loops.
Single-cell analyses revealed discontinuous transcription, or pulses,
rather than smooth, uniform expression. Determining the pulse mechanism
will require more investigation, but possibilities include reversible
chromatin modifications or variations in transcription factor binding.
Pulsing would provide exquisite
regulatory control, suggests Singer.
“A burst that makes a lot of message
might overshoot the amount of protein
needed. That can have deleterious
effects.” Better to turn on for a flicker,
wait to equilibrate, then cycle back on
if more protein is needed.
dscA is induced as Dictyostelium
GFP reveals a spot of dscA
form fruiting bodies. This increase
transcription (light blue).
did not come from stronger or more
frequent pulses, but rather from more pulsing cells. The new
recruits were often clustered, suggesting either that the cluster is
seeing the same developmental stimulus or that expressing cells
tell their neighbors to join in. Whether constitutive genes also
pulse remains to be determined.
J
eing supportive is just not enough for a keratin. Seyun Kim, Pauline Wong, and Pierre Coulombe (Johns Hopkins University, Baltimore, MD)
find that Keratin 17 (K17) also takes on signaling duties
for cell growth during the wound response.
Keratins are a family of structural proteins that
form intermediate filaments. Coulombe’s group had
found that embryos lacking K17 did not heal wounds
properly. They now find that these problems do not
stem from structural issues.
Wound-adjacent epithelial cells usually turn on
K17, grow larger, reduce adhesion, and polarize before migrating into the wound site. This marked cell
growth was noticeably stunted, however, in K17 mutants,
and the authors traced this defect to a block in mTORactivated protein synthesis.
At least one known contributor to mTOR activation,
the 14-3-3σ adaptor protein, was found with cytoplasmic K17 filaments in wild-type keratinocytes. In cells lacking K17, however, 14-3-3σ was mostly nuclear. In the
cytoplasm, 14-3-3σ might bring various components of
the mTOR pathway to K17, though the exact mechanism
of translational up-regulation by the keratin is unclear.
Coulombe has also found that keratins delay apoptosis in the hair follicle. Such signaling functions, he says,
“might be the missing link towards accounting for why
keratins exhibit such exquisite cell- and context-specific
regulation. Do we need 50 different keratins just for a
mechanical function? Probably not.”
Reference: Chubb, J.R., et al. 2006. Curr. Biol. 16:1018–1025.
Reference: Kim, S., et al. 2006. Nature. 441:362–365.
B
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Vi r u s s q u e e z e d s h u t
he DNA in a fully packed viral capsid squeezes a pressure sensor,
as revealed by images from Gabriel Lander, John Johnson (Scripps
Research Institute, La Jolla, CA), and colleagues. The embrace triggers
the end of DNA packaging.
Each particle of the p22 bacteriophage, a herpes cousin, contains a single copy
of its genome. The DNA is pumped as a long concatamer into one end of the
capsid through a portal of gp1 proteins. After one genome enters, the concatamer
is cleaved and the portal plugged shut. In gp1 mutants, too much DNA is let in, but
just how the wild-type portal senses full capacity was unknown.
Johnson’s group viewed the fully assembled portal using automated electron
microscopy, which identified items of interest systematically, thus retrieving ten times
as many images as in manual reconstructions. The higher resolution data revealed
features of the intact viral particle that were previously hidden, including the gp1
portal. “Everything we dreamed of seeing was staring us in the face,” said Johnson.
The portal had previously been seen as an isolated entity, but it looked much
different in the DNA-filled virion. A ring of DNA wrapped around the portal “sort
of like a belt,” said Johnson. “It looks like the DNA is squeezing the portal and
changing its conformation. This probably signals to the outside, ‘we are full of
DNA.’” The pumps then know to cut the DNA.
T
Reference: Lander, G.C., et al. 2006. Science. doi:10.1126/science.1127981.
JOHNSON/AAAS
CHUBB/ELSEVIER
[email protected]
In a full virion, a ring of DNA (green) squeezes
the gp1 portal (red) into a new conformation.
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